Kemppainen J A, Lane M V, Sar M, Wilson E M
Laboratory for Reproductive Biology, University of North Carolina, Chapel Hill 27599.
J Biol Chem. 1992 Jan 15;267(2):968-74.
Nuclear transport, phosphorylation, ligand binding, and degradation rate of the recombinant androgen receptor (AR) were analyzed in transfected COS cells in the presence of various steroids and antiandrogens. Transcriptional activation was assessed in CV1 cells by cotransfection with an androgen-responsive chloramphenicol acetyltransferase (CAT) reporter vector. Hormone binding specificity of recombinant AR was essentially identical to endogenous AR. AR localized in the nucleus in the presence of methyltrienolone (R1881, a synthetic androgen), dihydrotestosterone, testosterone, hydroxyflutamide, cyproterone acetate, estradiol, progesterone, and RU486. In the absence of hormone or with the antiandrogen, flutamide, AR remained largely in the cytoplasm with a perinuclear distribution. AR was degraded rapidly (t1/2 = 1 h) except in the presence of androgen (t1/2 = 6 h) which accounted for an apparent 2-4-fold androgen-induced increase in AR phosphorylation, indicating that AR phosphorylation was not enhanced by androgen. CAT activity was stimulated by R1881, dihydrotestosterone, testosterone, cyproterone acetate, estradiol, progesterone, and RU486 in a dose-dependent manner. The antiandrogens, flutamide and hydroxyflutamide, lacked agonist activity and inhibited R1881-induced activation of CAT and androgen stabilization of AR. Steroids and antiandrogens with moderate to low affinity for AR promoted both nuclear transport and transcriptional activation but only at high hormone concentrations. Hydroxyflutamide acted as a true antiandrogen since it lacked agonist activity and was an inhibitor of androgen-induced transcriptional activation.
在转染的COS细胞中，于各种类固醇和抗雄激素存在的情况下，分析了重组雄激素受体（AR）的核转运、磷酸化、配体结合及降解速率。通过与雄激素反应性氯霉素乙酰转移酶（CAT）报告载体共转染，在CV1细胞中评估转录激活情况。重组AR的激素结合特异性与内源性AR基本相同。在甲基三烯olone（R1881，一种合成雄激素）、二氢睾酮、睾酮、羟基氟他胺、醋酸环丙孕酮、雌二醇、孕酮和RU486存在时，AR定位于细胞核。在无激素或存在抗雄激素氟他胺时，AR主要保留在细胞质中，呈核周分布。除了在雄激素存在时（半衰期 = 6小时），AR降解迅速（半衰期 = 1小时），雄激素导致AR磷酸化明显增加2 - 4倍，表明雄激素并未增强AR磷酸化。R1881、二氢睾酮、睾酮、醋酸环丙孕酮、雌二醇、孕酮和RU486以剂量依赖性方式刺激CAT活性。抗雄激素氟他胺和羟基氟他胺缺乏激动剂活性，并抑制R1881诱导的CAT激活和AR的雄激素稳定作用。对AR具有中度至低度亲和力的类固醇和抗雄激素促进核转运和转录激活，但仅在高激素浓度时。羟基氟他胺是一种真正的抗雄激素，因为它缺乏激动剂活性，并且是雄激素诱导的转录激活的抑制剂。
